Nucleic Acids Isolation and DNA Analysis Facility

核心C的具体目标是： 1)LFS和WT患者的淋巴母细胞系和成纤维细胞培养物的建立。 2)从外周血分离DNA、RNA和血浆;以及从肿瘤和正常人分离DNA和RNA 研究个体的组织样本。这包括那些患有Wilms肿瘤的人， 潜在的p53突变携带者和这些个体的家庭成员。 3)通过对p53、WT 1和/或某些WT“途径”的PCR产物直接测序进行突变检测 儿童肉瘤激酶、肾母细胞瘤激酶和散发性肾母细胞瘤的基因 4)对WT患者进行Southern分析，以确定PCR测序无法检测到的WT 1缺失。 5)奇数p53错义突变的功能表征和潜在截短突变的鉴定 携带无义突变体和剪接突变体的个体细胞中的蛋白质 6)通过WAVE分析对项目4中确定的选定WT途径基因进行突变筛查 7)DNA、RNA和血浆样本的储存和分发（如需要） 8)维护数据库，详细说明样品处理、核酸分离和突变 分析.此数据库将用于更新由Core C管理的程序数据库。 9)序列和突变筛选输出的存档。 这些功能对于P01中所有项目的成功至关重要。项目1使用p53和WT 1数据 更好地描述癌症风险，并开发更好的癌症易感性遗传模型。的 关于p53突变状态的信息也将鉴定非p53家族， （项目1999年）2)负责易感性的其他基因，并指导肿瘤数据的解释 从突变小鼠中产生（项目3和5）。WT 1突变状态对于描绘 肾母细胞瘤和WT患者的分子亚群，并阐明 突变，改变和表达变化，我们以前已经确定在肾母细胞瘤（项目4）。 该核心提供样本处理、突变筛选和分析、功能分析、样本处理、基因组测序、基因 存储和数据存储对于P01的所有项目至关重要。

The specific goals for Core C are: 1) establishment of lymphoblastoid cell lines and fibroblast cultures on LFS and WT patients. 2) isolation of DNA, RNA, and plasma from peripheral blood; and DNA and RNA from tumor and normal tissue samples from study individuals. This includes those with Wilms tumors, those who are potential p53 mutation carriers, and family members of these individuals. 3) mutation detection by direct sequencing of PCR products for p53, WT1, and/or some WT "pathway" genes for childhood sarcoma kindreds, Wilms tumor kindreds, and sporadic Wilms tumors 4) Southern analysis of WT patients to identify WT1 deletions undetectable by PCR-based sequencing. 5) functional characterization of odd p53 missense mutations and identification of potentially truncated proteins in cells from individuals carrying nonsense and splicing mutants 6) mutation screening by WAVE analysis for selected WT pathway genes identified in Project 4 7) storage and distribution, as needed, of DNA, RNA, and plasma samples 8) maintenance of a database detailing sample handling, nucleic acids isolations, and mutational analysis. This database will be used to update the Program's database managed by Core C. 9) archiving of sequence and mutation screening output. These functions are critical to the success of all projects within the P01. Project 1 uses p53 and WT1 data to characterize better cancer risk and also to develop better genetic models for cancer predisposition. The information regarding p53 mutational status will also identify non-p53 families that will aid the identification (Proj. 2) of other genes responsible for predisposition and guide the interpretation of the tumor data generated from mutant mice (Projects 3 & 5). WT1 mutational status is critical for the delineation of the molecular subsets of Wilms tumors and WT patients and elucidating the biologic effect of the pattern of mutations, alterations, and expression changes we have previously identified in Wilms tumors (Project 4). This core provides sample processing, mutation screening and analyses, functional analyses, sample storage, and data storage critical for all projects of the P01.